HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Singh, C. R. Articles by Jagannath, C. Search for Related Content PubMed PubMed Citation Articles by Singh, C. R. Articles by Jagannath, C.

Processing and Presentation of a Mycobacterial Antigen 85B Epitope by Murine Macrophages Is Dependent on the Phagosomal Acquisition of Vacuolar Proton ATPase and In Situ Activation of Cathepsin D¹

Christopher R. Singh^*, Rachel A. Moulton^*, Lisa Y. Armitige^*, Akhil Bidani, Mark Snuggs, Subramanian Dhandayuthapani, Robert L. Hunter^* and Chinnaswamy Jagannath^2,*

^* Department of Pathology and Laboratory Medicine, Department of Pulmonary Medicine, and Department of Neurobiology, University of Texas Health Science Center, Houston, TX 77030; and Department of Microbiology, University of Texas Health Science Center, San Antonio, TX 78229

Mycobacterium tuberculosis (strain H37Rv) and bacillus Calmette-Guérin (BCG) vaccine inhibit phagosome maturation in macrophages and their effect on processing, and presentation of a secreted Ag85 complex B protein, Ag85B, by mouse macrophages was analyzed. Macrophages were infected with GFP-expressing mycobacterial strains and analyzed for in situ localization of vacuolar proton ATPase (v-ATPase) and cathepsin D (Cat D) using Western blot analysis and immunofluorescence. H37Rv and BCG phagosomes excluded the v-ATPase and maintained neutral pH while the attenuated H37Ra strain acquired v-ATPase and acidified. Mycobacterial phagosomes acquired Cat D, although strains BCG and H37Rv phagosomes contained the inactive 46-kDa form, whereas H37Ra phagosomes had the active 30-kDa form. Infected macrophages were overlaid with a T cell hybridoma specific for an Ag85B epitope complexed with MHC class II. Coincident with active Cat D, H37Ra-infected macrophages presented the epitope to T cells inducing IL-2, whereas H37Rv- and BCG-infected macrophages were less efficient in IL-2 induction. Bafilomycin inhibited the induction of macrophage-induced IL-2 from T cells indicating that v-ATPase was essential for macrophage processing of Ag85B. Furthermore, the small interfering RNA interference of Cat D synthesis resulted in a marked decrease in the levels of macrophage-induced IL-2. Thus, a v-ATPase-dependent phagosomal activation of Cat D was required for the generation of an Ag85B epitope by macrophages. Reduced processing of Ag85B by H37Rv- and BCG-infected macrophages suggests that phagosome maturation arrest interferes with the efficient processing of Ags in macrophages. Because Ag85B is immunodominant, this state may lead to a decreased ability of the wild-type as well as the BCG vaccine to induce protective immunity.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ This work was supported by Grant NI49534 from the National Institutes of Health.

² Address correspondence and reprint requests to Dr. Chinnaswamy Jagannath, Department of Pathology and Laboratory Medicine, Medical School Building 2.200, University of Texas Health Science Center, 6431 Fannin Street, Houston, TX 77030. E-mail address: Chinnaswamy.Jagannath{at}uth.tmc.edu

³ Abbreviations used in this paper: MTB, Mycobacterium tuberculosis; v-ATPase, vacuolar proton ATPase; Cat D, cathepsin D; BCG, M. bovis bacillus Calmette-Guérin; siRNA, small interfering RNA; MOI, multiplicity of infection; LAMP, lysosome-associated membrane protein; BMDM, bone marrow-derived macrophage; BPF, bodipy-pepstatin-FL.

This article has been cited by other articles:

				S. T. Chang, J. J. Linderman, and D. E. Kirschner Effect of Multiple Genetic Polymorphisms on Antigen Presentation and Susceptibility to Mycobacterium tuberculosis Infection Infect. Immun., July 1, 2008; 76(7): 3221 - 3232. [Abstract] [Full Text] [PDF]

				H. Soualhine, A.-E. Deghmane, J. Sun, K. Mak, A. Talal, Y. Av-Gay, and Z. Hmama Mycobacterium bovis Bacillus Calmette-Guerin Secreting Active Cathepsin S Stimulates Expression of Mature MHC Class II Molecules and Antigen Presentation in Human Macrophages J. Immunol., October 15, 2007; 179(8): 5137 - 5145. [Abstract] [Full Text] [PDF]

				J. A. MacGurn and J. S. Cox A Genetic Screen for Mycobacterium tuberculosis Mutants Defective for Phagosome Maturation Arrest Identifies Components of the ESX-1 Secretion System Infect. Immun., June 1, 2007; 75(6): 2668 - 2678. [Abstract] [Full Text] [PDF]